The present invention relates to a turbidimetric immunoassay for measuring the concentration of an antigen dissolved in a sample solution and more particularly to a turbidimetric immunoassay for measuring the concentration of albumin contained in urine, and an apparatus therefor.
Conventional methods for measuring the concentration of an antigen involve: adding, to a sample solution, an antibody that specifically binds to an antigen contained in the sample solution to produce an antigen-antibody complex by an antigen-antibody reaction, which makes the sample solution turbid; and determining the concentration of the antigen from the turbidity of the sample solution. The turbidity is measured by directing light to the sample solution and then detecting scattered light generated in the solution or transmitted light having passed through the solution. When the turbidity increases, the intensity of the scattered light also increases, but the intensity of the transmitted light decreases.
In the above-described method, the measurement is made in a range where the molar concentration of the antibody is greater than that of the antigen and a range where they are almost equivalent, more specifically, in prozone A through B (hereinafter may be referred to as A-B range) and zone of equivalence C (hereinafter may be referred to as C range) as shown in FIG. 17. In FIG. 17, the turbidity increases as the concentration of the antigen increases in the A-B range whereas the variation of the turbidity with the concentration of the antigen is small in the C range. In the postzone D, the turbidity decreases as the concentration of the antigen is increased, which is called prozone phenomenon. Accordingly, in the above method, the concentration of the antigen is determined by creating a calibration line in the A-B range of FIG. 17 and calculating it from a measured turbidity level based on the calibration curve.
As described above, the concentration of the antigen should be in the prozone A-B or the zone of equivalence C in order for the measurement to be made, and therefore the antibody concentration is required to be increased. In such case, sensitivity in a low concentration range is sometimes sacrificed. Moreover, in order to check that the concentration of the antigen is actually not in the postzone D (i.e. D range of FIG. 17), a step of diluting the sample solution or a step of adding additional amount of antibody has to be done. In the case of using a sample solution whose antigen concentration may be in the D range, two concentrations are obtained from measured turbidity levels. For this reason, it is necessary to check that the concentration of the antigen is not in the D range (see US2003-219910 A1 and US2003-100128 A1).